Differential expression of genetic information is required to direct the development of an undifferentiated, fertilized egg into a complex, multicellular organism. We are using Drosophila melanogaster as a ideal model system to study the mechanisms controlling developmentally regulated gene expression. The segmentation gene fushi tarazu (ftz) is expressed in a complex pattern during early embryogenesis. In the cellular blastoderm, ftz RNA is distributed in seven evenly spaced stripes that surround the embryo. We have identified two enhancers of the ftz gene that direct gene expression in ftz-like striped patterns when linked to a heterologous promoter. This study is focused on the ftz "proximal enhancer" which is the regulatory element that contains the most information for the correct spatial expression of ftz stripes. As part of this study, we have identified minimal cis-acting regulatory sequences within the proximal enhancer. A biochemical approach has been taken to identify the trans- acting factors that interact with the enhancer to direct the spatially and temporally restricted pattern of ftz expression during embryogenesis. This unbiased biochemical approach allows us to identify and purify transcription factors that positively and negatively regulate the dynamically evolving pattern of ftz expression. Specific mutations introduced into each of the identified binding motif will allow us to probe the in vivo relevance of identified binding sites. A protein interacting with an important regulatory region will be purified to homogeneity. Amino acid sequence data will be used to generate probes for gene cloning. In the future, the structure and function of the newly identified ftz regulatory protein(s) will be analyzed in vitro and in vivo. The long term goals of this work rest upon the premise that basic strategies controlling embryonic development have been conserved throughout the animal kingdom, as evidenced, for example, by the conservation of the homebox in developmentally regulated genes of a variety of species. The present studies will thus provide a framework for understanding the molecular mechanisms underlying determination and differentiation in Drosophila as well as in higher organisms.